This invention relates to methods for completing a cell culture medium with a lipophilic substance promoting growth of cells by weight or promoting the biosythesis of desired substances within the cells, and to nanoemulsions suitable in said methods.
Nanoemulsions, alternatively called nanoparticles, are composed of oil particles, the surfaces of which are occupied by an amphoteric emulsifier in aqueous dispersions. They can be prepared by mixing triglycerides or fatty acid esters in aqueous phase using a high pressure homogenizer (EP-B1-0 406 162xe2x80x94H. G. Weder).
So far, they were used for the manufacture of pharmaceutical and/or cosmetic preparations, and also for the preparation of nutrient solutions in which the nanoemulsions serve as the energy supplier in cell cultures (EP-B1-0 406 162xe2x80x94H. G. Weder).
The cell culture technique is a biological system having a very wide application. Controlled culturing of these cells is mainly used in immunology, in biotechnology, in toxicology, in gene technology, and in cell biology. In all said applications the interaction of substances with cells is of primary importance. The exchange of biochemical substances with the cells takes place in the culture medium which is composed of a large number of different substances. In order to create ideal conditions for the cells, in most cases blood serum is added to the culture medium. However, for many uses the complementation of said medium with blood serum shows severe disadvantages, since the blood serum has an undefined composition, is expensive, and may contain undesired components, such as viruses, prions and germs. Therefore, for many applications the creation of so-called serum-free media is of primary importance.
Working with cell cultures, a general difficulty is the application of test substances which are insoluble in water. If possible, lipophilic substances are dissolved in alcohol or dimethylsulfoxide, and then added to the culture medium. However, this results in undefined dispersions of said substances having a low bioavailability. The use of solubilizers, such as Tween 20(copyright), results in unstable and very toxic emulsions.
Therefore, no reliable and simple methods were so far available to solve the following problems:
Completion of cell culture media with lipophilic substances promoting cell growth;
Completion of cell culture media with lipophilic substances promoting biosynthesis of desired substances within the cells.
It is the primary object of the invention to solve the above mentioned problems by creating methods for delivering lipophilic substances to cell cultures, which lipophilic substances promote the growth of cells by weight or promote the biosythesis of desired substances within the cells, by using nanoemulsions as vehicle.
The foregoing and further objects, advantages and features will be apparent from the following specification.
To meet these and other objects, the invention provides the following methods, and the following dispersions suitable for use in these methods:
a method of completing a cell culture medium with a lipophilic substance promoting the growth of cells by weight, said method comprising the steps of: (a) preparing a dispersion consisting of a nanoemulsion containing the substance promoting the growth of cells, in aqueous phase; and (b) adding said dispersion to said cell culture;
a method of completing a cell culture medium with a lipophilic substance promoting the biosynthesis of desired substances within the cells, said method comprising the steps of: (a) preparing a dispersion consisting of a nanoemulsion containing the substance promoting said biosynthesis, in aqueous phase; and (b) adding said dispersion to said cell culture.
Preferably, said dispersions comprise as oily component a triglyceride of the fatty acids C 18:1 (oleic acid) and/or C 18:2 (linoleic acid), and comprise nanoemulsions comprising from 0.65 to 0.75 parts by weight of said amphoteric emulsifier for one part by weight of said oily component.